Characterization of the RelBbu Regulon in Borrelia burgdorferi Reveals Modulation of Glycerol Metabolism by (p)ppGpp

Abstract

The bacterial stringent response is triggered by deficiencies of available nutrients and other environmental stresses. It is mediated by 5'-triphosphate-guanosine-3'-diphosphate and 5'-diphosphate-guanosine-3'-diphosphate (collectively (p)ppGpp) and generates global changes in gene expression and metabolism that enable bacteria to adapt to and survive these challenges. Borrelia burgdorferi encounters multiple stressors in its cycling between ticks and mammals that could trigger the stringent response. We have previously shown that the B. burgdorferi stringent response is mediated by a single enzyme, RelBbu, with both (p)ppGpp synthase and hydrolase activities, and that a B. burgdorferi 297 relBbu null deletion mutant was defective in adapting to stationary phase, incapable of down-regulating synthesis of rRNA and could not infect mice. We have now used this deletion mutant and microarray analysis to identify genes comprising the rel regulon in B. burgdorferi cultured at 34°C, and found that transcription of genes involved in glycerol metabolism is induced by rel Bbu. Culture of the wild type parental strain, the rel Bbu deletion mutant and its complemented derivative at 34°C and 25°C in media containing glucose or glycerol as principal carbon sources revealed a growth defect in the mutant, most evident at the lower temperature. Transcriptional analysis of the glp operon for glycerol uptake and metabolism in these three strains confirmed that rel Bbu was necessary and sufficient to increase transcription of this operon in the presence of glycerol at both temperatures. These results confirm and extend previous findings regarding the stringent response in B. burgdorferi. They also demonstrate that the stringent response regulates glycerol metabolism in this organism and is likely crucial for its optimal growth in ticks.

Spermine and Spermidine Alter Gene Expression and Antigenic Profile of Borrelia burgdorferi

“…In combination with density-dependent signals, these data reflect the role of RpoS in Spd-treated spirochetes. Both rpoZ and rel Bbu (bb0198) have been implicated in the metabolic response observed when spirochetes are shifted from minimal growth medium to nutritionally replete conditions that reflect the conditions in the midgut of tick before and after a blood meal (14,15). The transcript levels of rel Bbu were significantly elevated in B. burgdorferi grown to low density in the presence of Spd, while rel Bbu was downregulated in the Spd-treated samples grown to high density (Fig.…”

“…Rel Bbu is a bifunctional synthetase/hydrolase controlling the levels of guanosine tetra/pentaphosphate [(p)ppGpp]. (p)ppGpp has been shown to play a role in the regulation of the stringent response during the survival of B. burgdorferi in the tick vector or during starvation (14,15). While temperature, growth phase, or pH of the medium did not affect the transcriptional level of rel Bbu , shifting the spirochetes from nutritionally starved medium to that supplemented with rabbit serum significantly induced its transcription (15).…”

“…A key in vitro growth component that allows the spirochetes to recover from starvation is rabbit serum, and this was reflected with increased transcription of rel Bbu (14,15). Importantly, polyamines affect the surface composition, notably porins, in many bacterial species.…”

Interaction of the Lyme disease spirochete with its tick vector

“…1). Modulating the levels of these second messengers in response to external signals alters the transcriptional landscape, as well as many other cellular processes, adapting the spirochete to the challenges of a dynamic and hostile environment in the tick midgut (Caimano et al, 2011; 2015; He et al, 2011; Kostick et al, 2011; Sultan et al, 2011; Bugrysheva et al, 2015; Drecktrah et al, 2015). Classically, (p)ppGpp synthesis is triggered by an accumulation of uncharged tRNAs during nutrient stress, but a shortage of carbon, iron, phosphate or fatty acids can also induce the stringent response (reviewed in Potrykus and Cashel, 2008; Dalebroux and Swanson, 2012; Hauryliuk et al, 2015).…”

“…The (p)ppGpp-dependent transcriptome, determined using a rel Bbu mutant, included a number of persistence-related genes induced by the stringent response, such as the glycerol uptake and metabolism ( glp ) operon (Bugrysheva et al, 2015; Drecktrah et al, 2015). The glp operon is induced in ticks and by glycerol in vitro ; survival of glp mutants is compromised in the tick, highlighting the importance of glycerol as a signal and carbon source in this part of the enzootic cycle (He et al, 2011; Pappas et al, 2011).…”